In the proposed continuation project, attempts will be made to study glucagon secretion under physiological and pathological conditions, including experimental diabetes mellitus. Islet of Langerhans will be isolated by a standardized method from normal rats, as well as from rats previously treated in various ways. The isolated islets will be perifused with buffers containing known or potential secretagogues and the glucagon and insulin content of the effluent media will be assayed by radioimmunoassay. In some instances, perfusion will begin with inhibitors of alpha-cell function. Experiments are designed to confirm increased glucagon secretion by islets of fasting, hypoglycemic and diabetic rats in response to stimuli such as free amino acids (ariginine), catecholamines and ionophores such as the divalent cation specific carrier A23187. Using these agents in vitro, it is hoped to obtain effects which will explain metabolic derangements associated with abnormal glucagon secretion in vivo. Attempts will also be made to produce anti-glucagon sera of sufficient specificity and activity that serum glucagon measurements can be made reliably on single animals.